Among the few hydrolytic enzymes active in the detoxication process, are imidase and the carboxylesterases and imidase. Several cytosolic carboxylesterases from rat liver have now been purified to homogeneity and characterized, two of which appear to be responsible for the hydrolysis of aspirin. Imidase, also a cytosolic protein, has now been purified to homogeneity from rat liver. As with all of the enzymes of detoxication, imidase is active with a broad range of substrates that include such cyclic imides as hydantoins; succinimide, glutimide and adipimide; dihydrouracil and dihydrothymidine; and the aliphatic imide, deacetylimide. Only the hydrolysis of the two pyrimidines to their respective ureido derivatives requires magnesium ions; with these two there is an observable reverse reaction. For the hydrolysis of uracil to yield N-carbamyl-beta-alanine, the equilibrium contrast was found to be 1 at pH 5.4. The enzyme exhibits stereoselectivity for one of the isomers of 2-methylhydantoin and stereospecificity for one of the isomers of 2-isopropylhydantoin and of 5-phenylsuccinimide. Imidase is a protein of about 240,000 daltons composed of four subunits of equal size. It is thought to catalyze one of the reactions in the degradative pathway of uridine and thymidine under the label of dihydropyrimidinase and hydropyrimidine hydrase; the term imidase reflects the broader potential of the enzyme that has now been purified.